This document describes a study examining stable isotope analyses of vegetation across Australian plant communities. It provides brief descriptions of nine Australian supersites that vary in climate and vegetation type. The study aims to examine spatial and seasonal variability in intrinsic water use efficiency across sites, determine the factors influencing differences in WUE within and among species, and identify plant functional types based on stable isotope analyses. The results could help understand niche separation and water use strategies of co-occurring species.

1. Stable isotope (13C) analyses of
vegetation across Australia
Derek Eamus
Rizwana Rumman
Tomek Wyczesany
with the co-operation of many Supersite Managers
across Australia

2. The Australian Supersite Network

3. Site descriptions
Veg Rainfall
(mm)
Temp
(oC)
Litchfield NT Savanna 1700 23 - 32
Far N Qld Simple notophyll vine forest 2000 + 23 - 29
Alice Springs NT Mulga (Acacia anuera) 300 14 - 30
SE Qld (peri-urban) a mix of dry eucalypt forest and wet
heath
1000 15 - 26
WA mosaic of temperate woodland,
heathland and mallee vegetation
245 11 - 25
Tumburumba wet eucalypt forest 900 5 - 20
Victoria dry eucalypt forest 675 7 - 18
Tasmania tall closed eucalypt forest 600 12 - 27
SA Undulating mallee woodland ; Calperum
Mallee
225 10 - 25

4. Aims of the study
• Examine seasonal and spatial variability in WUE
across Australia
• Examine niche separation of co-occurring species
• Determine the relative contributions of
differences in A and gs to differences in WUEi
across species and sites (needs Δ18O data of
cellulose)
• Determination of plant functional types based
upon stable isotope analyses

5. Extracting water (for deuterium and 18O
analyses) from soil and stem segments
using cryo-distillation

6. Picarro analysers for water and C
stable isotope analyses

7. • Why do plants contain less 13C in their leaves than the CO2 in
the air?
1. 13CO2 diffuses into leaves more slowly than 12CO2
2. Rubisco preferentially binds 12CO2
 δ13Cplant = δ13Catm - (diffusion effect + enzyme effect)
Discrimination

8. • The δ13C of C3 plants reflects how much stomatal conductance limits photosynthesis.
δ13Cplant, ‰
C3 plants C4 plants
Source: Cerling et al., 1997
less
stomatal
limitation
more
stomatal
limitation
ci ≈ ca; ci/ca=1
Less δ13C
More Δ
Smaller WUEi
ci << ca; ci/ca<1
More δ13C
Less Δ
Larger WUEi
Source: Werner et al, 2012, Cerling et al., 1997

9. • Farquhar, Ehleringer and Hubick (1989), proposed the
equation for carbon isotope discrimination:
 a = Diffusion effect (the rate of diffusion of 13CO2 across
stomatal pore from atmosphere to leaf is lower by a factor of
~4.4‰ than 12CO2)
a
i
c
c
aba )(
 b = Isotope effect caused by preference of Rubisco for 12CO2
over 13CO2 ~27 ‰

10. Ca
CO2
Ci
H2O
wi
wa
gc gw
• Carbon isotope discrimination is a measure of intrinsic water-
use-efficiency (WUEi)
 WUEi is the ratio of carbon assimilation to stomatal conductance
(A/gw)
 Photosynthesis, A = gc (ca-ci)
 WUEi = A/gw = gc (ca-ci)/gw
Source: Werner et al, 2012

12. NSW data (2 spp per site) + Alice Springs Supersite
+ Robson Creek + GDE River Red Gum
y = -0.0025x - 25.752
R² = 0.67
-34
-32
-30
-28
-26
-24
-22
-20
0 500 1000 1500 2000 2500
δ13C
Annual rainfall (mm)
Robson Creek
(FNQ)
River Red Gum, Alice
Springs - using GW –
thinks it is receiving 1800
mm of rainfall
Alice
Springs
New South
Wales data

13. Comparing our data with Miller et al. 2001 (NT
gradient) and Schulze et al. (1998; Qld gradient):
y = 0.002x + 18.21
R² = 0.670
y = 0.002x + 18.17
R² = 0.701
y = 1.079ln(x) + 12.61
R² = 0.630
12
14
16
18
20
22
24
26
28
30
0 500 1000 1500 2000 2500
Delta(%o)
Annual rainfall (mm)

14. Comparing our data with Stewart et al.
1995 (S Qld)
y = -0.0028x - 25.45
R² = 0.53
-32
-30
-28
-26
-24
-22
-20
0 200 400 600 800 1000 1200 1400 1600 1800
δ13C(%o)
Annual rainfall (mm)

15. What can we do with δ13C data?
• Can use δ13C to calculate Δ (discrimination)
since:
Δ = (-8- δ13C )/(1+ δ13C /1000)
• Can calculate WUEi and Ci/Ca from this:
WUEi = (Ca*(27- Δ ))/(1.6*(27-4.4))
Ci/Ca = (Δ -4.4)/(27-4.4)

16.  Using eddy covariance data available in the Supersites to
compare ecoWUE and stable isotope assessments of
WUEi
Recent studies show that:
 Ecosystem instantaneous water-use-efficiency ecoWUEinst
and ecosystem intrinsic water-use-efficiency ecoWUEi are
responsive to changes in soil moisture content and leaf-to-
air VPD (Vickers et al. 2012)
 an assessment at canopy-scales will enable the
comparison across contrasting communities (Campos et al. 2013)

17. WUEi across multiple sites
y = -0.0258x + 87.48
R² = 0.67
0
10
20
30
40
50
60
70
80
90
100
0 500 1000 1500 2000 2500
Intrinsicwater-use-efficiency
Mean annual rainfall (mm)

18. Example of application of stable isotope
analyses: Ecohydrological niche separation
Can we identify contrasting strategies for water-
use among co-existing plant species?
– Moreno-Gutierrez et al 2012: δ13C used to
disaggregate 10 co-occurring species along an
ecophysiological gradient
– Combined δ13C and Δ18O revealed a wide
spectrum of degree of stomatal regulation of A
– This correlated with WUEi and coexisting species
maintained their species specific isotopic niches
that reflect ecohydrological niche segregation

19. Niche separation ? Variation in δ13C within a site
across species is as large as across sites across a
rainfall gradient – data from FNQ
-35.00
-30.00
-25.00
-20.00
-15.00
-10.00
Alphitoniawhitei
Elaeocarpusgrandis
Flindersiabrayleyana
Acaciacelsa
Argyrodendronperlatum
Melicopeelleryana
Flindersiabrayleyana
Homolanthusnovoguineensis
Eiaeocarpusgrandis
Elaeocarpusgrandis
Alphitoniapetriei
Flindersiabrayleyana
Blepherocoryiainvoluaisula
Flindersiabrayleyana
Neolitseadealbata
Elaeocarpusgrandis
Castanospermumaustrale
Cardwelliasublimis
Plascospermuncoriaceum
castanosporaalphandii
Unknown
Flindersiabrayleyana
Elaeocarpusgrandis
Unknown
Aiphitoniapetriei
Aleuritesrockinghamensis
Alphitoniapetriei
Flindersiabrayleyana
Castanospermumaustrale
Flindersiabrayleyana
Flindersiabrayleyana
Aleuritesrockinghamensis
Acaciacelsa
Unknown
Alphitoniapetriei
Aistoniascholaris
Alstoniascholaris
Unknown.(Kaoripine?)
Alstoniascholaris
δ13C%o

20. Example of application of stable isotope
analyses: Plant Functional Type Analyses
Can we identify contrasting plant functional
types using Δ13C ?
– Werner and Maguas 2010: Compared Δ13C of
across 11 species of contrasting PFT
– Found pronounced and consistent variation in
Δ13C across PFTs – correlated with phenology, leaf
attributes (SLA, foliar N)
– Principal component analyses showed Δ13C to be
as good a predictor of PFT as using multiple leaf
traits.

21. Current data gaps
• Samples from different PFTs
• Wide selection of species across a site
• Δ18O of leaf cellulose to disaggregate effects of
gs from A

22. The END
Thanks for listening and
any questions?

23. Conclusions
• Do species matter?

24. Brief site descriptions
• Litchfield Savanna: tall open eucalypt savanna in a tropical wet-dry climate; mean
annual min/max temp of 23–32 oC; annual rainfall of 1700 mm
• FNQ Rainforest: a simple notophyll vine forest in a tropical wet climate; mean
annual min/max temp of 23–29 oC ; annual rainfall of 2000+ mm
• Alice Mulga: open Mulga woodland savanna in a semi-arid climate; mean annual
min/max temp of 14–30 oC; annual rainfall of 275 mm
• SEQ Peri-urban: a mix of dry eucalypt forest and wet heath in a subtropical
climate; mean annual min/max temp of 15–26 oC; annual rainfall of 1000 mm
• Great Western Woodland: a mosaic of temperate woodland, heathland and
mallee vegetation in a semi-arid climate; mean annual min/max temp of 11–25 oC;
annual rainfall of 245 mm

25. • Tumbarumba Wet Eucalypt: wet eucalypt forest in a temperate climate;
mean annual min/max temp of 5–20 oC; annual rainfall of 900 mm
• Victorian Dry Eucalypt: dry eucalypt forest is a temperate climate; mean
annual min/max temp of 7–18 oC; annual rainfall of 675 mm
• Warra Tall Eucalypt: a tall closed eucalypt forest in a temperate climate;
mean annual min/max temp of 12–27 oC; annual rainfall of 600 mm
• Cumberland Plain EucFace: remnant eucalypt woodland in a temperate
climate; mean annual min/max temp of 11–24 oC; annual rainfall 700 mm
• Calperum Mallee: an undulating mallee woodland in a semi-arid climate;
mean annual min/max temp of 10–25 oC; annual rainfall of 225 mm